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Innate immune signaling is essential for clearing pathogens and damaged cells and must be tightly regulated to avoid excessive inflammation or autoimmunity. Here, we found that the alternative splicing of exons derived from transposable elements is a key mechanism controlling immune signaling in human cells. By analyzing long-read transcriptome datasets, we identified numerous transposon exonization events predicted to generate functional protein variants of immune genes, including the type I interferon receptor IFNAR2. We demonstrated that the transposon-derived isoform of IFNAR2 is more highly expressed than the canonical isoform in almost all tissues and functions as a decoy receptor that potently inhibits interferon signaling, including in cells infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Our findings uncover a primate-specific axis controlling interferon signaling and show how a transposon exonization event can be co-opted for immune regulation., Competing Interests: Declaration of interests G.I.M.P. and E.B.C. have filed a provisional patent related to this work (PCT application no. PCT/US2023/066767)., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Replication of the complex retrovirus mouse mammary tumor virus (MMTV) is antagonized by murine Apobec3 (mA3), a member of the Apobec family of cytidine deaminases. We have shown that MMTV-encoded Rem protein inhibits proviral mutagenesis by the Apobec enzyme, activation-induced cytidine deaminase (AID) during viral replication in BALB/c mice. To further study the role of Rem in vivo, we have infected C57BL/6 (B6) mice with a superantigen-independent lymphomagenic strain of MMTV (TBLV-WT) or a mutant strain that is defective in Rem and its cleavage product Rem-CT (TBLV-SD). Compared to BALB/c, B6 mice were more susceptible to TBLV infection and tumorigenesis. Furthermore, unlike MMTV, TBLV induced T-cell tumors in B6 μMT mice, which lack membrane-bound IgM and conventional B-2 cells. At limiting viral doses, loss of Rem expression in TBLV-SD-infected B6 mice accelerated tumorigenesis compared to TBLV-WT in either wild-type B6 or AID-knockout mice. Unlike BALB/c results, high-throughput sequencing indicated that proviral G-to-A or C-to-T mutations were unchanged regardless of Rem expression in B6 tumors. However, knockout of both AID and mA3 reduced G-to-A mutations. Ex vivo stimulation showed higher levels of mA3 relative to AID in B6 compared to BALB/c splenocytes, and effects of agonists differed in the two strains. RNA-Seq revealed increased transcripts related to growth factor and cytokine signaling in TBLV-SD-induced tumors relative to TBLV-WT-induced tumors, consistent with another Rem function. Thus, Rem-mediated effects on tumorigenesis in B6 mice are independent of Apobec-mediated proviral hypermutation., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Byun et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The interplay between host factors and viral components impacts viral replication efficiency profoundly. Members of the cellular heterogeneous nuclear ribonucleoprotein family (hnRNPs) have been extensively studied as HIV-1 host dependency factors, but whether they play a role in innate immunity is currently unknown. This study aimed to identify hnRNPA0 as a type I interferon (IFN)-repressed host factor in HIV-1-infected cells. Knockdown of hnRNPA0, a situation that mirrors conditions under IFN stimulation, increased LTR activity, export of unspliced HIV-1 mRNA, viral particle production, and thus, increased infectivity. Conversely, hnRNPA0 overexpression primarily reduced plasmid-driven and integrated HIV-1 long terminal repeat (LTR) activity, significantly decreasing total viral mRNA and protein levels. In addition, high levels of hnRNPA0 significantly reduced the HIV-1 programmed ribosomal frameshifting efficiency, resulting in a shift in the HIV-1 p55/p15 ratio. The HIV-1 alternative splice site usage remained largely unaffected by altered hnRNPA0 levels suggesting that the synergistic inhibition of the LTR activity and viral mRNA transcription, as well as impaired ribosomal frameshifting efficiency, are critical factors for efficient HIV-1 replication regulated by hnRNPA0. The pleiotropic dose-dependent effects under high or low hnRNPA0 levels were further confirmed in HIV-1-infected Jurkat cells. Finally, our study revealed that hnRNPA0 levels in PBMCs were lower in therapy-naive HIV-1-infected individuals compared to healthy controls. Our findings highlight a significant role for hnRNPA0 in HIV-1 replication and suggest that its IFN-I-regulated expression levels are critical for viral fitness allowing replication in an antiviral environment.IMPORTANCERNA-binding proteins, in particular, heterogeneous nuclear ribonucleoproteins (hnRNPs), have been extensively studied. Some act as host dependency factors for HIV-1 since they are involved in multiple cellular gene expression processes. Our study revealed hnRNPA0 as an IFN-regulated host factor, that is differently expressed after IFN-I treatment in HIV-1 target cells and lower expressed in therapy-naïve HIV-1-infected individuals. Our findings demonstrate the significant pleiotropic role of hnRNPA0 in viral replication: In high concentrations, hnRNPA0 limits viral replication by negatively regulating Tat-LTR transcription, retaining unspliced mRNA in the nucleus, and significantly impairing programmed ribosomal frameshifting. Low hnRNPA0 levels as observed in IFN-treated THP-1 cells, particularly facilitate HIV LTR activity and unspliced mRNA export, suggesting a role in innate immunity in favor of HIV replication. Understanding the mode of action between hnRNPA0 and HIV-1 gene expression might help to identify novel therapeutically strategies against HIV-1 and other viruses., Competing Interests: The authors declare no conflict of interest.

Background: Endogenous retroelements (EREs), including human endogenous retroviruses (HERVs) and long interspersed nuclear elements (LINEs), comprise almost half of the human genome. Our previous studies of the interferome in the gut suggest potential mechanisms regarding how IFNb may drive HIV-1 gut pathogenesis. As ERE activity is suggested to partake in type 1 immune responses and is incredibly sensitive to viral infections, we sought to elucidate underlying interactions between ERE expression and gut dynamics in people living with HIV-1 (PLWH)., Methods: ERE expression profiles from bulk RNA sequencing of colon biopsies and PBMC were compared between a cohort of PLWH not on antiretroviral therapy (ART) and uninfected controls., Findings: 59 EREs were differentially expressed in the colon of PLWH when compared to uninfected controls (padj <0.05 and FC ≤ -1 or ≥ 1) [Wald's Test]. Of these 59, 12 EREs were downregulated in PLWH and 47 were upregulated. Colon expression of the ERE loci LTR19&#95;12p13.31 and L1FLnI&#95;1q23.1s showed significant correlations with certain gut immune cell subset frequencies in the colon. Furthermore L1FLnI&#95;1q23.1s showed a significant upregulation in peripheral blood mononuclear cells (PBMCs) of PLWH when compared to uninfected controls suggesting a common mechanism of differential ERE expression in the colon and PBMC., Interpretation: ERE activity has been largely understudied in genomic characterizations of human pathologies. We show that the activity of certain EREs in the colon of PLWH is deregulated, supporting our hypotheses that their underlying activity could function as (bio)markers and potential mediators of pathogenesis in HIV-1 reservoirs., Funding: US NIH grants NCI CA260691 (DFN) and NIAID UM1AI164559 (DFN)., Competing Interests: Declaration of interests All authors have no potential conflicts of interests to disclose., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Antiretroviral therapy (ART) transformed HIV from a life-threatening disease to a chronic condition. However, eliminating the virus remains an elusive therapy goal. For several decades, Friend virus (FV) infection serves as a murine model to study retrovirus immunity. Similar to HIV, FV persists at low levels in lymph nodes B cell follicles avoiding elimination by immune cells. Such immune-privileged reservoirs exclude cytotoxic T cells from entry. However, CXCR5+ T cells are permitted to traffic through germinal centers. This marker is predominantly expressed by CD4+ follicular helper T cells (Tfh). Therefore, we explored immunotherapy to induce cytotoxic Tfh, which are rarely found under physiological conditions. The TNF receptor family member CD137 was first identified as a promising target for cancer immunotherapy. We demonstrated that FV-infected mice treatment with αCD137 antibody resulted in an induction of the cytotoxic program in Tfh. The therapy significantly increased numbers of cytotoxic Tfh within B cell follicles and contributed to viral load reduction. Moreover, αCD137 antibody combined with ART delayed virus rebound upon treatment termination without disturbing the lymph node architecture or antibody responses. Thus, αCD137 antibody therapy might be a novel strategy to target the retroviral reservoir and an interesting approach for HIV cure research., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Malyshkina et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Innate immune signaling is essential for clearing pathogens and damaged cells, and must be tightly regulated to avoid excessive inflammation or autoimmunity. Here, we found that the alternative splicing of exons derived from transposable elements is a key mechanism controlling immune signaling in human cells. By analyzing long-read transcriptome datasets, we identified numerous transposon exonization events predicted to generate functional protein variants of immune genes, including the type I interferon receptor IFNAR2. We demonstrated that the transposon-derived isoform of IFNAR2 is more highly expressed than the canonical isoform in almost all tissues, and functions as a decoy receptor that potently inhibits interferon signaling including in cells infected with SARS-CoV-2. Our findings uncover a primate-specific axis controlling interferon signaling and show how a transposon exonization event can be co-opted for immune regulation., Competing Interests: Declaration of Interests: We have a patent application related to this work (PCT Application No. PCT/US2023/066767). Authors declare that they have no further competing interests.

We evaluated clinical and socioeconomic burdens of respiratory disease in banana farm workers in Guatemala. We offered all eligible workers enrollment during June 15-December 30, 2020, and annually, then tracked them for influenza-like illnesses (ILI) through self-reporting to study nurses, sentinel surveillance at health posts, and absenteeism. Workers who had ILI submitted nasopharyngeal swab specimens for testing for influenza virus, respiratory syncytial virus, and SARS-CoV-2, then completed surveys at days 0, 7, and 28. Through October 10, 2021, a total of 1,833 workers reported 169 ILIs (12.0 cases/100 person-years), and 43 (25.4%) were laboratory-confirmed infections with SARS-CoV-2 (3.1 cases/100 person-years). Workers who had SARS-CoV-2‒positive ILIs reported more frequent anosmia, dysgeusia, difficulty concentrating, and irritability and worse clinical and well-being severity scores than workers who had test result‒negative ILIs. Workers who had positive results also had greater absenteeism and lost income. These results support prioritization of farm workers in Guatemala for COVID-19 vaccination.

Human papillomavirus (HPV) infection is a causative agent of multiple human cancers, including cervical and head and neck cancers. In these HPV-positive tumors, somatic mutations are caused by aberrant activation of DNA mutators such as members of the apolipoprotein B messenger RNA-editing enzyme catalytic polypeptide-like 3 (APOBEC3) family of cytidine deaminases. APOBEC3 proteins are most notable for their restriction of various viruses, including anti-HPV activity. However, the potential role of APOBEC3 proteins in HPV-induced cancer progression has recently garnered significant attention. Ongoing research stems from the observations that elevated APOBEC3 expression is driven by HPV oncogene expression and that APOBEC3 activity is likely a significant contributor to somatic mutagenesis in HPV-positive cancers. This review focuses on recent advances in the study of APOBEC3 proteins and their roles in HPV infection and HPV-driven oncogenesis. Further, we discuss critical gaps and unanswered questions in our understanding of APOBEC3 in virus-associated cancers.

Humoral immune perturbations contribute to pathogenic outcomes in persons with HIV-1 infection (PWH). Gut barrier dysfunction in PWH is associated with microbial translocation and alterations in microbial communities (dysbiosis), and IgA, the most abundant immunoglobulin (Ig) isotype in the gut, is involved in gut homeostasis by interacting with the microbiome. We determined the impact of HIV-1 infection on the antibody repertoire in the gastrointestinal tract by comparing Ig gene utilization and somatic hypermutation (SHM) in colon biopsies from PWH ( n  = 19) versus age and sex-matched controls ( n  = 13). We correlated these Ig parameters with clinical, immunological, microbiome and virological data. Gene signatures of enhanced B cell activation were accompanied by skewed frequencies of multiple Ig Variable genes in PWH. PWH showed decreased frequencies of SHM in IgA and possibly IgG, with a substantial loss of highly mutated IgA sequences. The decline in IgA SHM in PWH correlated with gut CD4 + T cell loss and inversely correlated with mucosal inflammation and microbial translocation. Diminished gut IgA SHM in PWH was driven by transversion mutations at A or T deoxynucleotides, suggesting a defect not at the AID/APOBEC3 deamination step but at later stages of IgA SHM. These results expand our understanding of humoral immune perturbations in PWH that could have important implications in understanding mucosal immune defects in individuals with chronic HIV-1 infection. IMPORTANCE The gut is a major site of early HIV-1 replication and pathogenesis. Extensive CD4 + T cell depletion in this compartment results in a compromised epithelial barrier that facilitates the translocation of microbes into the underlying lamina propria and systemic circulation, resulting in chronic immune activation. To date, the consequences of microbial translocation on the mucosal humoral immune response (or vice versa) remains poorly integrated into the panoply of mucosal immune defects in PWH. We utilized next-generation sequencing approaches to profile the Ab repertoire and ascertain frequencies of somatic hypermutation in colon biopsies from antiretroviral therapy-naive PWH versus controls. Our findings identify perturbations in the Ab repertoire of PWH that could contribute to development or maintenance of dysbiosis. Moreover, IgA mutations significantly decreased in PWH and this was associated with adverse clinical outcomes. These data may provide insight into the mechanisms underlying impaired Ab-dependent gut homeostasis during chronic HIV-1 infection.

The emergence of SARS-CoV-2 variants with enhanced transmissibility, pathogenesis, and resistance to vaccines presents urgent challenges for curbing the COVID-19 pandemic. While Spike mutations that enhance virus infectivity or neutralizing antibody evasion may drive the emergence of these novel variants, studies documenting a critical role for interferon responses in the early control of SARS-CoV-2 infection, combined with the presence of viral genes that limit these responses, suggest that interferons may also influence SARS-CoV-2 evolution. Here, we compared the potency of 17 different human interferons against multiple viral lineages sampled during the course of the global outbreak, including ancestral and five major variants of concern that include the B.1.1.7 (alpha), B.1.351 (beta), P.1 (gamma), B.1.617.2 (delta), and B.1.1.529 (omicron) lineages. Our data reveal that relative to ancestral isolates, SARS-CoV-2 variants of concern exhibited increased interferon resistance, suggesting that evasion of innate immunity may be a significant, ongoing driving force for SARS-CoV-2 evolution. These findings have implications for the increased transmissibility and/or lethality of emerging variants and highlight the interferon subtypes that may be most successful in the treatment of early infections.

Essential agricultural workers work under occupational conditions that may increase the risk of SARS-CoV-2 exposure and transmission. Data from an agricultural worker cohort in Guatemala, and anti-SARS-CoV-2 nucleocapsid IgG (anti-N IgG) testing were used to estimate past infections and analyze risk factors associated with seropositivity at enrollment and association with SARS-CoV-2 infection. The stability of neutralizing antibody (NAb) responses were assessed in a subset of participants. The adjusted relative risk (aRR) for seroprevalence at enrollment was estimated accounting for correlations within worksites. At enrollment, 616 (46.2%) of 1334 (93.2%) participants had anti-N IgG results indicating prior SARS-CoV-2 infection. A cough ≤ 10 days prior to enrollment (aRR = 1.28, 95% CI: 1.13−1.46) and working as a packer (aRR = 2.00, 95% CI: 1.67−2.38) or packing manager within the plants (aRR = 1.82, 95% CI: 1.36−2.43) were associated with increased risk of seropositivity. COVID-19 incidence density among seronegative workers was 2.3/100 Person-Years (P-Y), higher than seropositive workers (0.4/100 P-Y). Most workers with follow-up NAb testing (65/77, 84%) exhibited a 95% average decrease in NAb titers in <6 months. While participants seropositive at baseline were less likely to experience a symptomatic SARS-CoV-2 infection during follow-up, NAb titers rapidly waned, underscoring the need for multipronged COVID-19 prevention strategies in the workplace, including vaccination.

Background: Primary Sjögren syndrome (pSS) is a chronic autoimmune disease with its main target being exocrine glands, and is the connective tissue disease more frequently associated with other autoimmune diseases. The aim of this study was to assess the frequency of another autoimmune rheumatic disease (ARD) developed in primary Sjögren syndrome (pSS) patients and to describe it's clinical, serological and histologic characteristics., Materials and Methods: This is a retrospective cohort study. Data of patients with pSS diagnosis (American-European criteria 2002), included in the GESSAR database (Grupo de Estudio Síndrome de Sjögren, Sociedad Argentina de Reumatología) were analyzed. The development of a second ARD was registered during the follow up., Results: 681 patients were included, 94.8% female. The mean age was 54 (SD 14) years and mean age at diagnosis of 50 (SD 13) years. The mean follow-up was 4.7 (SD 4.9) years; 30 patients (4.41%, CI 95%: 3.1-5.7) developed a second ARD during the follow up, incidence rate was 9.1/1000 patients-year (IR 95%: 5.8-12.4/1000 patients-year), the most frequent being rheumatoid arthritis (RA). 96% out of these 30 patients had xerophthalmia, 86.2% xerostomia, 92% positive Schirmer test, 88.24% positive Rosa Bengala test, lisamine green or Ocular Staining Score, 81.2% positive unstimulated salivary flow, 82.1% Ro(+) and 33.33% La(+). Minor salivary gland biopsy had been performed in 14 of the 30 patients, 12 with positive results. There were no statistically significant differences respect baseline characteristics when comparing the patients who developed another ARD to the ones that did not., Conclusions: Of all the patients analyzed, 4.4% presented another ARD during their follow-up. It is important to be aware of this, to make an early and proper diagnosis and treatment of our patients., (© 2022. The Author(s).)

The impacts of interferon (IFN) signaling on COVID-19 pathology are multiple, with both protective and harmful effects being documented. We report here a multiomics investigation of systemic IFN signaling in hospitalized COVID-19 patients, defining the multiomics biosignatures associated with varying levels of 12 different type I, II, and III IFNs. The antiviral transcriptional response in circulating immune cells is strongly associated with a specific subset of IFNs, most prominently IFNA2 and IFNG. In contrast, proteomics signatures indicative of endothelial damage and platelet activation associate with high levels of IFNB1 and IFNA6. Seroconversion and time since hospitalization associate with a significant decrease in a specific subset of IFNs. Additionally, differential IFN subtype production is linked to distinct constellations of circulating myeloid and lymphoid immune cell types. Each IFN has a unique metabolic signature, with IFNG being the most associated with activation of the kynurenine pathway. IFNs also show differential relationships with clinical markers of poor prognosis and disease severity. For example, whereas IFNG has the strongest association with C-reactive protein and other immune markers of poor prognosis, IFNB1 associates with increased neutrophil to lymphocyte ratio, a marker of late severe disease. Altogether, these results reveal specialized IFN action in COVID-19, with potential diagnostic and therapeutic implications., Competing Interests: Competing interest statement: J.M.E. serves on the COVID Development Advisory Board for Eli Lilly., (Copyright © 2022 the Author(s). Published by PNAS.)

We evaluated the clinical and socioeconomic burdens of respiratory disease in a cohort of Guatemalan banana plantation workers. All eligible workers were offered enrollment from June 15-December 30, 2020, and annually, then followed for influenza-like illnesses (ILI) through: 1) self-reporting to study nurses, 2) sentinel surveillance at health posts, and 3) absenteeism. Workers with ILI submitted nasopharyngeal swabs for influenza, RSV, and SARS-CoV-2 testing, then completed surveys at days 0, 7, and 28. Through October 10, 2021, 1,833 workers developed 169 ILIs (12.0/100 person-years) and 43 (25.4%) of these ILIs were laboratory-confirmed SARS-CoV-2 (3.1/100 person-years). Workers with SARS-CoV-2-positive ILI reported more anosmia (p<0.01), dysgeusia (p<0.01), difficulty concentrating (p=0.01), and irritability (p=0.01), and greater clinical and well-being severity scores (Flu-iiQ) than test-negative ILIs; they also had greater absenteeism (p<0.01) and lost income (median US$127.1, p<0.01). These results support the prioritization of Guatemalan farm workers for COVID-19 vaccination.

SAMHD1 is a potent HIV-1 restriction factor that blocks reverse transcription in monocytes, dendritic cells and resting CD4 + T cells by decreasing intracellular dNTP pools. However, SAMHD1 may diminish innate immune sensing and Ag presentation, resulting in a weaker adaptive immune response. To date, the role of SAMHD1 on antiretroviral immunity remains unclear, as mouse SAMHD1 had no impact on murine retrovirus replication in prior in vivo studies. Here, we show that SAMHD1 significantly inhibits acute Friend retrovirus infection in mice. Pretreatment with LPS, a significant driver of inflammation during HIV-1 infection, further unmasked a role for SAMHD1 in influencing immune responses. LPS treatment in vivo doubled the intracellular dNTP levels in immune compartments of SAMHD1 knockout but not wild-type mice. SAMHD1 knockout mice exhibited higher plasma infectious viremia and proviral DNA loads than wild-type mice at 7 d postinfection (dpi), and proviral loads inversely correlated with a stronger CD8 + T cell response. SAMHD1 deficiency was also associated with weaker NK, CD4 + T and CD8 + T cell responses by 14 dpi and weaker neutralizing Ab responses by 28 dpi. Intriguingly, SAMHD1 influenced these cell-mediated immune (14 dpi) and neutralizing Ab (28 dpi) responses in male but not female mice. Our findings formally demonstrate SAMHD1 as an antiretroviral factor in vivo that could promote adaptive immune responses in a sex-dependent manner. The requirement for LPS to unravel the SAMHD1 immunological phenotype suggests that comorbidities associated with a "leaky" gut barrier may influence the antiviral function of SAMHD1 in vivo., (Copyright © 2022 by The American Association of Immunologists, Inc.)

The dried-tube specimen (DTS) procedure was used to develop the COVID-19 serology control panel (CSCP). The DTS offers the benefit of shipping materials without a cold chain, allowing for greater access without deterioration of material integrity. Samples in the panel were sourced from COVID-19 convalescent persons from March to May 2020. The immunoglobulin subtypes (total Ig, IgM, and IgG) and their respective reactivity to severe acute respiratory syndrome coronavirus 2 nucleocapsid, spike, and receptor-binding domain antigens of the samples were delineated and compared with the WHO International Standard to elucidate the exact binding antibody units of each CSCP sample and ensure the CSCP provides adequate reactivity for different types of serological test platforms. We distribute the CSCP as a kit with five coded tubes to laboratories around the world to be used to compare test kits for external quality assurance, for harmonizing laboratory testing, and for use as training materials for laboratory workers.

Chronic HIV-1 infection results in the sustained disruption of gut homeostasis culminating in alterations in microbial communities (dysbiosis) and increased microbial translocation. Major questions remain on how interactions between translocating microbes and gut immune cells impact HIV-1-associated gut pathogenesis. We previously reported that in vitro exposure of human gut cells to enteric commensal bacteria upregulated the serine protease and cytotoxic marker Granzyme B (GZB) in CD4 T cells, and GZB expression was further increased in HIV-1-infected CD4 T cells. To determine if these in vitro findings extend in vivo , we evaluated the frequencies of GZB + CD4 T cells in colon biopsies and peripheral blood of untreated, chronically infected people with HIV-1 (PWH). Colon and blood GZB + CD4 T cells were found at significantly higher frequencies in PWH. Colon, but not blood, GZB + CD4 T cell frequencies were associated with gut and systemic T cell activation and Prevotella species abundance. In vitro , commensal bacteria upregulated GZB more readily in gut versus blood or tonsil-derived CD4 T cells, particularly in inflammatory T helper 17 cells. Bacteria-induced GZB expression in gut CD4 T cells required the presence of accessory cells, the IL-2 pathway and in part, MHC Class II. Overall, we demonstrate that GZB + CD4 T cells are prevalent in the colon during chronic HIV-1 infection and may emerge following interactions with translocated bacteria in an IL-2 and MHC Class II-dependent manner. Associations between GZB + CD4 T cells, dysbiosis and T cell activation suggest that GZB + CD4 T cells may contribute to gut HIV-1 pathogenesis.

The emergence of SARS-CoV-2 variants with enhanced transmissibility, pathogenesis and resistance to vaccines presents urgent challenges for curbing the COVID-19 pandemic. While Spike mutations that enhance virus infectivity or neutralizing antibody evasion may drive the emergence of these novel variants, studies documenting a critical role for interferon responses in the early control of SARS-CoV-2 infection, combined with the presence of viral genes that limit these responses, suggest that interferons may also influence SARS-CoV-2 evolution. Here, we compared the potency of 17 different human interferons against multiple viral lineages sampled during the course of the global outbreak, including ancestral and four major variants of concern. Our data reveal increased interferon resistance in emerging SARS-CoV-2 variants, suggesting that evasion of innate immunity may be a significant, ongoing driving force for SARS-CoV-2 evolution. These findings have implications for the increased lethality of emerging variants and highlight the interferon subtypes that may be most successful in the treatment of early infections.

Severe coronavirus disease 2019 (COVID-19) has been associated with T cell lymphopenia, but no causal effect of T cell deficiency on disease severity has been established. To investigate the specific role of T cells in recovery from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections, we studied rhesus macaques that were depleted of either CD4 + , CD8 + , or both T cell subsets prior to infection. Peak virus loads were similar in all groups, but the resolution of virus in the T cell-depleted animals was slightly delayed compared to that in controls. The T cell-depleted groups developed virus-neutralizing antibody responses and class switched to IgG. When reinfected 6 weeks later, the T cell-depleted animals showed anamnestic immune responses characterized by rapid induction of high-titer virus-neutralizing antibodies, faster control of virus loads, and reduced clinical signs. These results indicate that while T cells play a role in the recovery of rhesus macaques from acute SARS-CoV-2 infections, their depletion does not induce severe disease, and T cells do not account for the natural resistance of rhesus macaques to severe COVID-19. Neither primed CD4 + nor CD8 + T cells appeared critical for immunoglobulin class switching, the development of immunological memory, or protection from a second infection. IMPORTANCE Patients with severe COVID-19 often have decreased numbers of T cells, a cell type important in fighting most viral infections. However, it is not known whether the loss of T cells contributes to severe COVID-19 or is a consequence of it. We studied rhesus macaques, which develop only mild COVID-19, similar to most humans. Experimental depletion of T cells slightly prolonged their clearance of virus, but there was no increase in disease severity. Furthermore, they were able to develop protection from a second infection and produced antibodies capable of neutralizing the virus. They also developed immunological memory, which allows a much stronger and more rapid response upon a second infection. These results suggest that T cells are not critical for recovery from acute SARS-CoV-2 infections in this model and point toward B cell responses and antibodies as the essential mediators of protection from re-exposure.

Group 3 innate lymphoid cells (ILC3s) in the gut mucosa have long been thought to be noncytotoxic lymphocytes that are critical for homeostasis of intestinal epithelial cells through secretion of IL-22. Recent work using human tonsillar cells demonstrated that ILC3s exposed to exogenous inflammatory cytokines for a long period of time acquired expression of granzyme B, suggesting that under pathological conditions ILC3s may become cytotoxic. We hypothesized that inflammation associated with bacterial exposure might trigger granzyme B expression in gut ILC3s. To test this, we exposed human colon lamina propria mononuclear cells to a panel of enteric bacteria. We found that the Gram-negative commensal and pathogenic bacteria induced granzyme B expression in a subset of ILC3s that were distinct from IL-22-producing ILC3s. A fraction of granzyme B + ILC3s coexpressed the cytolytic protein perforin. Granzyme B expression was mediated, in part, by IL-15 produced upon exposure to bacteria. ILC3s coexpressing all three IL-15R subunits (IL15Rα/β/γ) increased following bacterial stimulation, potentially allowing for cis presentation of IL-15 during bacterial exposure. Additionally, a large frequency of colonic myeloid dendritic cells expressed IL-15Rα, implicating myeloid dendritic cells in trans presentation of IL-15 to ILC3s. Tonsillar ILC3s minimally expressed granzyme B when exposed to the same bacteria or to rIL-15. Overall, these data establish the novel, to our knowledge, finding that human colonic ILC3s can express granzyme B in response to a subset of enteric bacteria through a process mediated by IL-15. These observations raise new questions about the multifunctional role of human gut ILC3s., (Copyright © 2021 by The American Association of Immunologists, Inc.)

In this study, we present the photosynthetic oxygen (O 2 ) supply to mammalian cells within a volumetric extracellular matrix-like construct, whereby a three-dimensional (3D)-bioprinted fugitive pattern encapsulating unicellular green algae, Chlamydomonas reinhardtii ( C. reinhardtii ), served as a natural photosynthetic O 2 -generator. The presence of bioprinted C. reinhardtii enhanced the viability and functionality of mammalian cells while reducing the hypoxic conditions within the tissues. We were able to subsequently endothelialize the hollow perfusable microchannels formed after enzymatic removal of the bioprinted C. reinhardtii -laden patterns from the matrices following the initial oxygenation period, to obtain biologically relevant vascularized mammalian tissue constructs. The feasibility of co-culture of C. reinhardtii with human cells, the printability and the enzymatic degradability of the fugitive bioink, as well as the exploration of C. reinhardtii as a natural, eco-friendly, cost-effective, and sustainable source of O 2 would likely promote the development of engineered tissues, tissue models, and food for various applications., Competing Interests: AUTHOR CONTRIBUTIONS S.M. designed and performed the experiments, collected and analyzed the data, and prepared the manuscript; J.A., C.C., A.G.R., D.H, C.D., E.C., M.D.R., D.B., M.L.S, W.L., F.C., and G.Y. performed the experiments and revised the manuscript; Y.S.Z. conceptualized, designed, and supported the study, and prepared the manuscript. DECLARATION OF INTERESTS The authors declare no competing interest.

The Type I Interferons (IFN-Is) are innate antiviral cytokines that include 12 different IFNα subtypes and IFNβ that signal through the IFN-I receptor (IFNAR), inducing hundreds of IFN-stimulated genes (ISGs) that comprise the 'interferome'. Quantitative differences in IFNAR binding correlate with antiviral activity, but whether IFN-Is exhibit qualitative differences remains controversial. Moreover, the IFN-I response is protective during acute HIV-1 infection, but likely pathogenic during the chronic stages. To gain a deeper understanding of the IFN-I response, we compared the interferomes of IFNα subtypes dominantly-expressed in HIV-1-exposed plasmacytoid dendritic cells (1, 2, 5, 8 and 14) and IFNβ in the earliest cellular targets of HIV-1 infection. Primary gut CD4 T cells from 3 donors were treated for 18 hours ex vivo with individual IFN-Is normalized for IFNAR signaling strength. Of 1,969 IFN-regulated genes, 246 'core ISGs' were induced by all IFN-Is tested. However, many IFN-regulated genes were not shared between the IFNα subtypes despite similar induction of canonical antiviral ISGs such as ISG15, RSAD2 and MX1, formally demonstrating qualitative differences between the IFNα subtypes. Notably, IFNβ induced a broader interferome than the individual IFNα subtypes. Since IFNβ, and not IFNα, is upregulated during chronic HIV-1 infection in the gut, we compared core ISGs and IFNβ-specific ISGs from colon pinch biopsies of HIV-1-uninfected (n = 13) versus age- and gender-matched, antiretroviral-therapy naïve persons with HIV-1 (PWH; n = 19). Core ISGs linked to inflammation, T cell activation and immune exhaustion were elevated in PWH, positively correlated with plasma lipopolysaccharide (LPS) levels and gut IFNβ levels, and negatively correlated with gut CD4 T cell frequencies. In sharp contrast, IFNβ-specific ISGs linked to protein translation and anti-inflammatory responses were significantly downregulated in PWH, negatively correlated with gut IFNβ and LPS, and positively correlated with plasma IL6 and gut CD4 T cell frequencies. Our findings reveal qualitative differences in interferome induction by diverse IFN-Is and suggest potential mechanisms for how IFNβ may drive HIV-1 pathogenesis in the gut., Competing Interests: The authors have declared that no competing interests exist.

Peripheral tolerance is essential for silencing weakly autoreactive B cells that have escaped central tolerance, but it is unclear why these potentially pathogenic B cells are retained rather than being eliminated entirely. Release from peripheral tolerance restraint can occur under certain circumstances (i.e., strong TLR stimulus), that are present during infection. In this regard, we hypothesized that autoreactive B cells could function as a reserve population that can be activated to contribute to the humoral immune response, particularly with pathogens, such as HIV-1, that exploit immune tolerance to avoid host defense. In this study, we identify a population of autoreactive B cells with the potential to neutralize HIV-1 and experimentally release them from the functional restrictions of peripheral tolerance. We have previously identified murine monoclonal antibodies that displayed autoreactivity against histone H2A and neutralized HIV-1 in vitro . Here, we identify additional H2A-reactive IgM monoclonal antibodies and demonstrate that they are both autoreactive and polyreactive with self and foreign antigens and are able to neutralize multiple clades of tier 2 HIV-1. Flow cytometric analysis of H2A-reactive B cells in naïve wildtype mice revealed that these B cells are present in peripheral B cell populations and we further document that murine H2A-reactive B cells are restrained by peripheral tolerance mechanisms. Specifically, we show endogenous H2A-reactive B cells display increased expression of the inhibitory mediators CD5 and phosphatase and tensin homolog (PTEN) phosphatase and fail to mobilize calcium upon immunoreceptor stimulation; all characterized markers of anergy. Moreover, we show that toll-like receptor stimulation or provision of CD4 T cell help induces the in vitro production of H2A-reactive antibodies, breaking tolerance. Thus, we have identified a novel poly/autoreactive B cell population that has the potential to neutralize HIV-1 but is silenced by immune tolerance., (Copyright © 2020 Agazio, Cimons, Shotts, Guo, Santiago, Pelanda and Torres.)

The innate immune system is normally programmed for immediate but transient upregulation in response to invading pathogens, and interferon (IFN)-stimulated gene (ISG) activation is a central feature. In contrast, chronic innate immune system activation is typically associated with autoimmunity and a broad array of autoinflammatory diseases that include the interferonopathies. Here, we studied retroviral susceptibility in a transgenic mouse model with lifelong innate immune system hyperactivation. The mice transgenically express low levels of a picornaviral RNA-dependent RNA polymerase (RdRP), which synthesizes double-stranded RNAs that are sensed by melanoma differentiation-associated protein 5 (MDA5) to trigger constitutive upregulation of many ISGs. However, in striking counterpoint to the paradigm established by numerous human and murine examples of ISG hyperactivation, including constitutive MDA5 activation, they lack autoinflammatory sequelae. RdRP-transgenic mice (RdRP mice) resist infection and disease caused by several pathogenic RNA and DNA viruses. However, retroviruses are sensed through other mechanisms, persist in the host, and have distinctive replication and immunity-evading properties. We infected RdRP mice and wild-type (WT) mice with various doses of a pathogenic retrovirus (Friend virus) and assessed immune parameters and disease at 1, 4, and 8 weeks. Compared to WT mice, RdRP mice had significantly reduced splenomegaly, viral loads, and infection of multiple target cell types in the spleen and the bone marrow. During chronic infection, RdRP mice had 2.35 ± 0.66 log 10 lower circulating viral RNA than WT. Protection required ongoing type I IFN signaling. The results show that the reconfigured RdRP mouse innate immune system substantially reduced retroviral replication, set point, and pathogenesis. IMPORTANCE Immune control of retroviruses is notoriously difficult, a fundamental problem that has been most clinically consequential with the HIV-1 pandemic. As humans expand further into previously uninhabited areas, the likelihood of new zoonotic retroviral exposures increases. The role of the innate immune system, including ISGs, in controlling retroviral infections is currently an area of intensive study. This work provides evidence that a primed innate immune system is an effective defense against retroviral pathogenesis, resulting in reduced viral replication and burden of disease outcomes. RdRP mice also had considerably lower Friend retrovirus (FV) viremia. The results could have implications for harnessing ISG responses to reduce transmission or control pathogenesis of human retroviral pathogens., (Copyright © 2020 American Society for Microbiology.)

Gut CD4 T cells are major targets of HIV-1 and are massively depleted early during infection. To better understand the mechanisms governing HIV-1-mediated CD4 T cell death, we developed the physiologically-relevant Lamina Propria Aggregate Culture (LPAC) model. The LPAC model is ideal for studying CD4 T cell death induced by clinically-relevant Transmitted/Founder (TF) HIV-1 strains and is also suitable for studying how enteric microbes and soluble factors ( e.g. , Type I Interferons) impact LP CD4 T cell death and function. Here, we detail the protocol to establish LP CD4 T cell infection using a process of spinoculation, the subsequent evaluation of infection levels using multicolor flow cytometry and the determination of overall LP CD4 T cell death using absolute LP CD4 T cell counts. We also describe the preparation of virus stocks of Transmitted/Founder (TF) HIV-1 infectious molecular clones that were successfully used in the LPAC model., Competing Interests: Competing interestsThe authors declare no conflicts of interest., (Copyright © 2020 The Authors; exclusive licensee Bio-protocol LLC.)

The innate immune response induced by type I interferons (IFNs) plays a critical role in the establishment of HIV infection. IFNs are induced early in HIV infection and trigger an antiviral defense program by signaling through the IFNα/β receptor (IFNAR), which consists of two subunits, IFNAR1 and IFNAR2. Changes in IFNAR expression in HIV target cells, as well as other immune cells, could therefore have important consequences for initial HIV spread. It was previously reported that IFNAR2 expression is increased in peripheral blood CD4+ CXCR4+ T cells of HIV+ patients compared to HIV uninfected controls, suggesting that HIV infection may alter the IFN responsiveness of target cells. However, the earliest immune cells affected by HIV in vivo reside in the gut-associated lymphoid tissue (GALT). To date, it remains unknown if IFNAR expression is altered in GALT immune cells in the context of HIV infection and exposure to IFNs, including the 12 IFNα subtypes. Here, we analyzed the expression of surface bound and soluble IFNAR2 on Lamina propria mononuclear cells (LPMCs) isolated from the GALT of HIV- individuals and in plasma samples of HIV+ patients. IFNAR2 expression varied between different T cells, B cells and natural killer cells, but was not altered following HIV infection. Furthermore, expression of the soluble IFNAR2a isoform was not changed in HIV+ patients compared to healthy donors, nor in LPMCs after HIV-1 infection ex vivo. Even though the 12 human IFNα subtypes trigger different biological responses and vary in their affinity to both receptor subunits, stimulation of LPMCs with different recombinant IFNα subtypes did not result in any significant changes in IFNAR2 surface expression. Our data suggests that potential changes in the IFN responsiveness of mucosal immune cells during HIV infection are unlikely dictated by changes in IFNAR2 expression., Competing Interests: The authors have declared that no competing interests exist.

Objective: To adapt the EULAR Activity Index for primary Sjögren's syndrome (ESSDAI) to the Argentine population., Methods: observational, cross-sectional study that included patients in a period of ten months. Three Argentine rheumatologists adapted and translated to Spanish the original version in English and the final version was translated back into English by a research associate whose mother language was English. In order to estimate the constructive validity of the index, the visual analogous scale (VAS) of disease activity was used by experts. A subgroup of patients attended a second visit in order to evaluate test-retest reliability., Results: 51 patients were included, 49 (96.1%) were female, the median age was 58 ((interquartile range (IQR): 49-69)). The median global VAS was 10 (IQR: 4-22.25) and the median total ESSDAI score was 5 (IQR: 3-9). The correlation between the global VAS and the total ESSDAI score of the scale was 0.79. The intraclass correlation coefficient was 0.67 (95% CI: 0.32-0.92) for the total score and 0.98 (95% CI: 0.92-0.995) for the global VAS. The results of the correlation coefficient between the VAS and the scale for each domain were: constitutional symptoms: 0.46; lymphadenopathy: 0.76; glandular: 0.78; joint: 0.61; skin: 1; respiratory: 0.83; renal: 1; muscular:- (no patient had myositis); peripheral nervous system: 0.72; central nervous system: 0.67; hematological: 0.96; biomarkers: 0.86., Conclusion: The results of this study showed that the ESSDAI is a reliable and valid index for this pSS argentinian population.

Clinical administration of Interferon α (IFNα) resulted in limited therapeutic success against some viral infections. Immune modulation of CD8 + T cell responses during IFNα therapy is believed to play a pivotal role in promoting viral clearance. However, these clinical studies primarily focused on IFNα subtype 2. To date, the immunomodulatory roles of the remaining 10-13 IFNα subtypes remains poorly understood, thereby precluding assessments of their potential for more effective treatments. Here, we report that virus-specific CD8 + T cell responses were influenced to various extents by individual IFNα subtypes. IFNα4, 6, and 9 had the strongest effects on CD8 + T cells, including antiproliferative effects, improved cytokine production and cytotoxicity. Interestingly, augmented cytokine responses were dependent on IFNα subtype stimulation of dendritic cells (DCs), while antiproliferative effects and cytotoxicity were mediated by IFNAR signaling in either CD8 + T cells or DCs. Thus, precise modulation of virus-specific CD8 + T cell responses may be feasible for specific antiviral immunotherapies through careful selection and administration of individual IFNα subtypes., (Copyright © 2019 Dickow, Francois, Kaiserling, Malyshkina, Drexler, Westendorf, Lang, Santiago, Dittmer and Sutter.)

Approximately 4.4% of the human genome is comprised of endogenous retroviral sequences, a record of an evolutionary battle between man and retroviruses. Much of what we know about viral immunity comes from studies using mouse models. Experiments using the Friend virus (FV) model have been particularly informative in defining highly complex anti-retroviral mechanisms of the intrinsic, innate and adaptive arms of immunity. FV studies have unraveled fundamental principles about how the immune system controls both acute and chronic viral infections. They led to a more complete understanding of retroviral immunity that begins with cellular sensing, production of type I interferons, and the induction of intrinsic restriction factors. Novel mechanisms have been revealed, which demonstrate that these earliest responses affect not only virus replication, but also subsequent innate and adaptive immunity. This review on FV immunity not only surveys the complex host responses to a retroviral infection from acute infection to chronicity, but also highlights the many feedback mechanisms that regulate and counter-regulate the various arms of the immune system. In addition, the discovery of molecular mechanisms of immunity in this model have led to therapeutic interventions with implications for HIV cure and vaccine development., (Published by Oxford University Press on behalf of FEMS 2019.)

The aim of this study was to evaluate the reliability of the outcome measures in rheumatology (OMERACT) definitions for ultrasound (US) elementary lesions in gout through an image reading exercise. Images from patients with gout (static images and videos) were collected. As an initial step, we carried out a image reading exercise within the experts of the Pan-American League of Associations for Rheumatology (PANLAR) US Study Group (n = 16). The following step consisted in a web-based exercise with the participation of larger number of sonographers (n = 63) from different centers. Images were rated evaluating the presence/absence of any US elementary lesion. Inter- and intra-reader reliabilities were analyzed using kappa coefficients. Participants were stratified according to their level of experience. In the first exercise, inter-reader kappa values were 0.45 for aggregates, 0.57 for tophus, 0.69 for erosions, and 0.90 for double contour (DC). Intra-reader kappa values were 0.86, 0.76, 0.80, and 0.90, respectively. The web-based exercise showed inter-reader kappa values for aggregates, tophus, erosions, and DC of 0.42, 0.49, 0.69, and 0.79, respectively. The intra-reader kappa values were 0.62, 0.69, 0.77, and 0.85, respectively. Reliability was not influenced by the sonographer's level of experience. The reliability of the new OMERACT US definitions for elementary lesions in gout ranged from moderate to excellent, depending on the type of lesion.

Innate lymphoid cells (ILCs) are a diverse family of cells that play critical roles in mucosal immunity. One subset of the ILC family, Group 3 ILCs (ILC3s), has been shown to aid in gut homeostasis through the production of IL-22. IL-22 promotes gut homeostasis through its functional effect on the epithelial barrier. When gut epithelial barrier integrity is compromised, such as in Human Immunodeficiency Virus (HIV) infection and inflammatory bowel disease (IBD), microbes from the gut lumen translocate into the lamina propria, inducing a multitude of potentially pathogenic immune responses. In murine models of bacterial infection, there is evidence that bacteria can induce pro-inflammatory IFNγ production in ILC3s. However, the impact of diverse translocating bacteria, particularly commensal bacteria, in dictating IFNγ versus IL-22 production by human gut ILC3s remains unclear. Here, we utilized an in vitro human lamina propria mononuclear cell (LPMC) model to evaluate ILC3 cytokine production in response to a panel of enteric Gram-positive and Gram-negative commensal and pathogenic bacteria and determined potential mechanisms by which these cytokine responses were induced. The percentages of IL-22-producing ILC3s, but not IFNγ-producing ILC3s, were significantly increased after LPMC exposure to both Gram-positive and Gram-negative commensal or pathogenic bacterial stimuli. Stimulation of IL-22 production from ILC3s was not through direct recognition of bacterial antigen by ILC3s, but rather required the help of accessory cells within the LPMC population. CD11c+ myeloid dendritic cells generated IL-23 and IL-1β in response to enteric bacteria and contributed to ILC3 production of IL-22. Furthermore, ligation of the natural cytotoxicity receptor NKp44 on ILC3s in response to bacteria stimulation also significantly increased the percentage of IL-22-producing ILC3s. Overall, these data demonstrate that human gut microbiota, including commensal bacteria, indirectly modulate colonic ILC3 function to induce IL-22, but additional signals are likely required to induce IFNγ production by colonic ILC3s in the setting of inflammation and microbial translocation.

Lymph- and blood-borne retroviruses exploit CD169/Siglec-1-mediated capture by subcapsular sinus and marginal zone metallophilic macrophages for trans-infection of permissive lymphocytes. However, the impact of CD169-mediated virus capture on retrovirus dissemination and pathogenesis in vivo is unknown. In a murine model of the splenomegaly-inducing retrovirus Friend virus complex (FVC) infection, we find that while CD169 promoted draining lymph node infection, it limited systemic spread to the spleen. At the spleen, CD169-expressing macrophages captured incoming blood-borne retroviruses and limited their spread to the erythroblasts in the red pulp where FVC manifests its pathogenesis. CD169-mediated retroviral capture activated conventional dendritic cells 1 (cDC1s) and promoted cytotoxic CD8 + T cell responses, resulting in efficient clearing of FVC-infected cells. Accordingly, CD169 blockade led to higher viral loads and accelerated death in susceptible mouse strains. Thus, CD169 plays a protective role during FVC pathogenesis by reducing viral dissemination to erythroblasts and eliciting an effective cytotoxic T lymphocyte response via cDC1s., (Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.)

Objective(s): Type I interferon (IFN-I) responses confer both protective and pathogenic effects in persistent virus infections. IFN-I diversity, stage of infection and tissue compartment may account for this dichotomy. The gut is a major site of early HIV-1 replication and microbial translocation, but the nature of the IFN-I response in this compartment remains unclear., Design: Samples were obtained from two IRB-approved cross-sectional studies. The first study included individuals with chronic, untreated HIV-1 infection (n = 24) and age/sex-balanced uninfected controls (n = 14). The second study included antiretroviral-treated, HIV-1-infected individuals (n = 15) and uninfected controls (n = 15)., Methods: The expression of 12 IFNα subtypes, IFNβ and antiviral IFN-stimulated genes (ISGs) were quantified in peripheral blood mononuclear cells (PBMCs) and colon biopsies using real-time PCR and next-generation sequencing. In untreated HIV-1-infected individuals, associations between IFN-I responses and gut HIV-1 RNA levels as well as previously established measures of colonic and systemic immunological indices were determined., Results: IFNα1, IFNα2, IFNα4, IFNα5 and IFNα8 were upregulated in PBMCs during untreated chronic HIV-1 infection, but IFNβ was undetectable. By contrast, IFNβ was upregulated and all IFNα subtypes were downregulated in gut tissue. Gut ISG levels positively correlated with gut HIV-1 RNA and immune activation, microbial translocation and inflammation markers. Gut IFN-I responses were not significantly different between HIV-1-infected individuals on antiretroviral treatment and uninfected controls., Conclusion: The IFN-I response is compartmentalized during chronic untreated HIV-1 infection, with IFNβ being more predominant in the gut. Gut IFN-I responses are associated with immunopathogenesis, and viral replication is likely a major driver of this response.

Objective: There are different imaging techniques to assess the parotid glands (i.e., sialography, salivary gland scintigraphy) in patients with Sjögren syndrome (SS). However, their use is limited by the invasive character or high cost. Ultrasound (US) is gaining interest by rheumatologists as a complementary diagnostic tool for SS. To date, there is an increasing body of evidence supporting its sensitivity in the assessment of salivary glands in SS. The aim of our study was to analyze the potential role of US as a diagnostic and prognostic tool in SS and to discuss existing evidence to support its application use., Methods: A systematic search was performed in the electronic database PubMed, using the following search terms: (salivary glands OR parotid glands OR submandibular glands) AND Sjögren's syndrome AND (ultrasonography OR ultrasound OR sonography). Titles, abstracts, and full reports were systematically screened., Results: The results of the studies analyzed in this review show encouraging results in terms of accuracy, validity, and diagnostic value, which leads us to believe that in the future US could become the reference imaging tool to assess SS. The studies include a small cohort of patients, and there is no standardized approach in terms of US techniques for the assessment of salivary glands., Conclusions: Ultrasound of major salivary glands is a useful tool for diagnosis, prognostic evaluation, and response to treatment in SS. The use of this imaging technology is still under development, and more multicentric studies are needed to validate this tool.

Sterile alpha motif and HD-domain-containing protein 1 (SAMHD1) blocks replication of retroviruses and certain DNA viruses by reducing the intracellular dNTP pool. SAMHD1 has been suggested to down-regulate IFN and inflammatory responses to viral infections, although the functions and mechanisms of SAMHD1 in modulating innate immunity remain unclear. Here, we show that SAMHD1 suppresses the innate immune responses to viral infections and inflammatory stimuli by inhibiting nuclear factor-κB (NF-κB) activation and type I interferon (IFN-I) induction. Compared with control cells, infection of SAMHD1-silenced human monocytic cells or primary macrophages with Sendai virus (SeV) or HIV-1, or treatment with inflammatory stimuli, induces significantly higher levels of NF-κB activation and IFN-I induction. Exogenous SAMHD1 expression in cells or SAMHD1 reconstitution in knockout cells suppresses NF-κB activation and IFN-I induction by SeV infection or inflammatory stimuli. Mechanistically, SAMHD1 inhibits NF-κB activation by interacting with NF-κB1/2 and reducing phosphorylation of the NF-κB inhibitory protein IκBα. SAMHD1 also interacts with the inhibitor-κB kinase ε (IKKε) and IFN regulatory factor 7 (IRF7), leading to the suppression of the IFN-I induction pathway by reducing IKKε-mediated IRF7 phosphorylation. Interactions of endogenous SAMHD1 with NF-κB and IFN-I pathway proteins were validated in human monocytic cells and primary macrophages. Comparing splenocytes from SAMHD1 knockout and heterozygous mice, we further confirmed SAMHD1-mediated suppression of NF-κB activation, suggesting an evolutionarily conserved property of SAMHD1. Our findings reveal functions of SAMHD1 in down-regulating innate immune responses to viral infections and inflammatory stimuli, highlighting the importance of SAMHD1 in modulating antiviral immunity., Competing Interests: The authors declare no conflict of interest.

APOBEC3 (A3) mutation signatures have been observed in a variety of human cancer genomes, including those of cervical and head and neck cancers caused by human papillomavirus (HPV) infection. However, the driving forces that promote off-target A3 activity remain mostly unclear. Here, we report a mechanism for the dramatic increase of A3A protein levels in HPV-positive keratinocytes. We show that expression of the viral protein E7 from high-risk HPVs, but not E7 from low-risk HPVs, significantly prolongs the cellular half-life of A3A protein in human keratinocytes and HPV-positive cancer cell lines. We have mapped several residues within the cullin 2 (CUL2) binding motif of HPV16 E7 as being important for mediating A3A protein stabilization. Furthermore, we provide direct evidence that both A3A and HPV16 E7 interact with CUL2, suggesting that the E7-CUL2 complex formed during HPV infection may regulate A3A protein levels in the cell. Using an in vitro cytidine deaminase assay, we show that E7-stabilized A3A remains catalytically active. Taken together, our findings suggest that the HPV oncoprotein E7 dysregulates endogenous A3A protein levels and thus provides novel mechanistic insight into cellular triggers of A3 mutations in HPV-positive cancers. IMPORTANCE Human papillomavirus (HPV) is causally associated with over 5% of all human malignancies. Several recent studies have shown that a subset of cancers, including HPV-positive head and neck and cervical cancers, have distinct mutational signatures potentially caused by members of the APOBEC3 cytidine deaminase family. However, the mechanism that induces APOBEC3 activity in cancer cells is poorly understood. Here, we report that the HPV oncoprotein E7 stabilizes the APOBEC3A (A3A) protein in human keratinocytes by inhibiting ubiquitin-dependent protein degradation in a cullin-dependent manner. Interestingly, the HPV E7-stabilized A3A protein maintains its deaminase activity. These findings provide a new insight into cancer mutagenesis enhanced by virus-induced A3A protein stabilization., (Copyright © 2018 Westrich et al.)

Follicular regulatory T (TFR) cells are a subset of CD4 + T cells in secondary lymphoid follicles. TFR cells were previously included in the follicular helper T (TFH) cell subset, which consists of cells that are highly permissive to HIV-1. The permissivity of TFR cells to HIV-1 is unknown. We find that TFR cells are more permissive than TFH cells to R5-tropic HIV-1 ex vivo TFR cells expressed more CCR5 and CD4 and supported higher frequencies of viral fusion. Differences in Ki67 expression correlated with HIV-1 replication. Inhibiting cellular proliferation reduced Ki67 expression and HIV-1 replication. Lymph node cells from untreated HIV-infected individuals revealed that TFR cells harbored the highest concentrations of HIV-1 RNA and highest levels of Ki67 expression. These data demonstrate that TFR cells are highly permissive to R5-tropic HIV-1 both ex vivo and in vivo This is likely related to elevated CCR5 levels combined with a heightened proliferative state and suggests that TFR cells contribute to persistent R5-tropic HIV-1 replication in vivo IMPORTANCE In chronic, untreated HIV-1 infection, viral replication is concentrated in secondary lymphoid follicles. Within secondary lymphoid follicles, follicular helper T (TFH) cells have previously been shown to be highly permissive to HIV-1. Recently, another subset of T cells in secondary lymphoid follicles was described, follicular regulatory T (TFR) cells. These cells share some phenotypic characteristics with TFH cells, and studies that showed that TFH cells are highly permissive to HIV-1 included TFR cells in their definition of TFH cells. The permissivity of TFR cells to HIV-1 has not previously been described. Here, we show that TFR cells are highly permissive to HIV-1 both ex vivo and in vivo The expression of Ki67, a marker of proliferative capacity, is predictive of expression of viral proteins, and downregulating Ki67 leads to concurrent decreases in expression of viral proteins. Our study provides new insight into HIV-1 replication and a potential new cell type to target for future treatment., (Copyright © 2017 American Society for Microbiology.)

A subset of characterized HIV-1 broadly neutralizing antibodies (bnAbs) are polyreactive with additional specificities for self-antigens and it has been proposed immunological tolerance may present a barrier to their participation in protective humoral immunity. We address this hypothesis by immunizing autoimmune-prone mice with HIV-1 Envelope (Env) and characterizing the primary antibody response for HIV-1 neutralization. We find autoimmune mice generate neutralizing antibody responses to tier 2 HIV-1 strains with alum treatment alone in the absence of Env. Importantly, experimentally breaching immunological tolerance in wild-type mice also leads to the production of tier 2 HIV-1-neutralizing antibodies, which increase in breadth and potency following Env immunization. In both genetically prone and experimentally induced mouse models of autoimmunity, increased serum levels of IgM anti-histone H2A autoantibodies significantly correlated with tier 2 HIV-1 neutralization, and anti-H2A antibody clones were found to neutralize HIV-1. These data demonstrate that breaching peripheral tolerance permits a cross-reactive HIV-1 autoantibody response able to neutralize HIV-1., (© 2017 Schroeder et al.)

Background: APOBEC3/Rfv3 restricts acute Friend retrovirus (FV) infection and promotes virus-specific neutralizing antibody (NAb) responses. Classical Rfv3 studies utilized FV stocks containing lactate-dehydrogenase elevating virus (LDV), a potent type I interferon inducer. Previously, we showed that APOBEC3 is required for the anti-FV activity of exogenous IFN-alpha treatment. Thus, type I interferon receptor (IFNAR) signaling may be required for the APOBEC3/Rfv3 response., Results: To test if the APOBEC3/Rfv3 response is dependent on type I IFN signaling, we infected IFNAR knockout versus IFNAR/APOBEC3 double-knockout mice with FV/LDV or LDV-free FV, and evaluated acute FV infection and subsequent NAb titers. We show that LDV co-infection and type I IFN signaling are not required for innate APOBEC3-mediated restriction. By contrast, removal of LDV and/or type I IFN signaling abrogated the APOBEC3-dependent NAb response., Conclusions: APOBEC3 can restrict retroviruses in a type I IFN-independent manner in vivo. By contrast, the ability of APOBEC3 to promote NAb responses is type I IFN-dependent. These findings reveal novel insights on the interplay between type I IFNs and APOBEC3 in vivo that may have implications for augmenting antiretroviral NAb responses.

Transient suppression of B cell function often accompanies acute viral infection. However, the molecular signaling circuitry that enforces this hyporesponsiveness is undefined. In this study, experiments identify up-regulation of the inositol phosphatase PTEN (phosphatase and tensin homolog) as primarily responsible for defects in B lymphocyte migration and antibody responses that accompany acute viral infection. B cells from mice acutely infected with gammaherpesvirus 68 are defective in BCR- and CXCR4-mediated activation of the PI3K pathway, and this, we show, is associated with increased PTEN expression. This viral infection-induced PTEN overexpression appears responsible for the suppression of antibody responses observed in infected mice because PTEN deficiency or expression of a constitutively active PI3K rescued function of B cells in infected mice. Conversely, induced overexpression of PTEN in B cells in uninfected mice led to suppression of antibody responses. Finally, we demonstrate that PTEN up-regulation is a common mechanism by which infection induces suppression of antibody responses. Collectively, these findings identify a novel role for PTEN during infection and identify regulation of the PI3K pathway, a mechanism previously shown to silence autoreactive B cells, as a key physiological target to control antibody responses., (© 2017 Getahun et al.)

Global transcriptome studies can help pinpoint key cellular pathways exploited by viruses to replicate and cause pathogenesis. Previous data showed that laboratory-adapted HIV-1 triggers significant gene expression changes in CD4+ T cell lines and mitogen-activated CD4+ T cells from peripheral blood. However, HIV-1 primarily targets mucosal compartments during acute infection in vivo. Moreover, early HIV-1 infection causes extensive depletion of CD4+ T cells in the gastrointestinal tract that herald persistent inflammation due to the translocation of enteric microbes to the systemic circulation. Here, we profiled the transcriptome of primary intestinal CD4+ T cells infected ex vivo with transmitted/founder (TF) HIV-1. Infections were performed in the presence or absence of Prevotella stercorea, a gut microbe enriched in the mucosa of HIV-1-infected individuals that enhanced both TF HIV-1 replication and CD4+ T cell death ex vivo. In the absence of bacteria, HIV-1 triggered a cellular shutdown response involving the downregulation of HIV-1 reactome genes, while perturbing genes linked to OX40, PPAR and FOXO3 signaling. However, in the presence of bacteria, HIV-1 did not perturb these gene sets or pathways. Instead, HIV-1 enhanced granzyme expression and Th17 cell function, inhibited G1/S cell cycle checkpoint genes and triggered downstream cell death pathways in microbe-exposed gut CD4+ T cells. To gain insights on these differential effects, we profiled the gene expression landscape of HIV-1-uninfected gut CD4+ T cells exposed to bacteria. Microbial exposure upregulated genes involved in cellular proliferation, MAPK activation, Th17 cell differentiation and type I interferon signaling. Our findings reveal that microbial exposure influenced how HIV-1 altered the gut CD4+ T cell transcriptome, with potential consequences for HIV-1 susceptibility, cell survival and inflammation. The HIV-1- and microbe-altered pathways unraveled here may serve as a molecular blueprint to gain basic insights in mucosal HIV-1 pathogenesis.

Objective: Gut microbial translocation is a major driving force behind chronic immune activation during HIV-1 infection. HIV-1-related intestinal dysbiosis, including increases in mucosa-associated pathobionts, may influence microbial translocation and contribute to mucosal and systemic inflammation. Thus, it is critical to understand the mechanisms by which gut microbes and their metabolic products, such as butyrate, influence immune cell function during HIV-1 infection., Design: A cross-sectional study was performed to compare the relative abundance of butyrate-producing bacterial (BPB) species in colonic biopsies and stool of untreated, chronic HIV-1-infected (n = 18) and HIV-1-uninfected (n = 14) study participants. The effect of exogenously added butyrate on gut T-cell activation and HIV-1 infection was evaluated using an ex-vivo human intestinal cell culture model., Methods: Species were identified in 16S ribosomal RNA sequence datasets. Ex-vivo isolated lamina propria mononuclear cells were infected with C-C chemokine receptor type 5-tropic HIV-1Bal, cultured with enteric gram-negative bacteria and a range of butyrate doses, and lamina propria T-cell activation and HIV-1 infection levels measured., Results: Relative abundance of total BPB and specifically of Roseburia intestinalis, were lower in colonic mucosa of HIV-1-infected versus HIV-1-uninfected individuals. In HIV-1-infected study participants, R. intestinalis relative abundance inversely correlated with systemic indicators of microbial translocation, immune activation, and vascular inflammation. Exogenous butyrate suppressed enteric gram-negative bacteria-driven lamina propria T-cell activation and HIV-1 infection levels in vitro., Conclusion: Reductions in mucosal butyrate from diminished colonic BPB may exacerbate pathobiont-driven gut T-cell activation and HIV replication, thereby contributing to HIV-associated mucosal pathogenesis.

Knowledge of the processes that underlie IgG subclass switching could inform strategies designed to counteract infections and autoimmunity. Here we show that TLR7 ligands induce subsets of memory CD4 and CD8 T cells to secrete interferon γ (IFNγ) in the absence of antigen receptor stimulation. In turn, TLR ligation and IFNγ cause B cells to express the transcription factor, T-bet, and to switch immunoglobulin production to IgG2a/c. Absence of TLR7 in T cells leads to the impaired T-bet expression in B cells and subsequent inefficient IgG2a isotype switching both in vitro and during the infection with Friend virus in vivo. Our results reveal a surprising mechanism of antiviral IgG subclass switching through T-cell intrinsic TLR7/IL-12 signaling., Competing Interests: The authors have declared that no competing interests exist.

Old World monkey (OWM) and hominid APOBEC3Aproteins exhibit differential restriction activities against lentiviruses and DNA viruses. Human APOBEC3A(hA3A)has weak restriction activity against HIV-1Δvifbut is efficiently restricted by an artificially generated chimeric from mandrills (mndA3A/G). We show that a chimeric hA3Acontaining the "WVS" insertion (hA3A[(27)WVS(29)]) conferred potent HIV-1restriction activity. Analysis of each amino acid of the "WVS" motif show that the length and not necessarily the charge or hydrophobicity of the amino acids accounted for restriction activity. Our results suggest that hA3A[(27)WVS(29)]restricts HIV-1at the level of reverse transcription in target cells. Finally, our results suggest that insertion of "WVS" into hA3Amodestly reduces restriction of adeno-associated virus 2(AAV-2)while insertion of the AC Loop1region of the mndA3A/G into hA3A abolished AAV-2 restriction, strengthening the role of this molecular interface in the functional evolution of primate A3A., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Unlabelled: Although all 12 subtypes of human interferon alpha (IFN-α) bind the same receptor, recent results have demonstrated that they elicit unique host responses and display distinct efficacies in the control of different viral infections. The IFN-α2 subtype is currently in HIV-1 clinical trials, but it has not consistently reduced viral loads in HIV-1 patients and is not the most effective subtype against HIV-1 in vitro We now demonstrate in humanized mice that, when delivered at the same high clinical dose, the human IFN-α14 subtype has very potent anti-HIV-1 activity whereas IFN-α2 does not. In both postexposure prophylaxis and treatment of acute infections, IFN-α14, but not IFN-α2, significantly suppressed HIV-1 replication and proviral loads. Furthermore, HIV-1-induced immune hyperactivation, which is a prognosticator of disease progression, was reduced by IFN-α14 but not IFN-α2. Whereas ineffective IFN-α2 therapy was associated with CD8(+) T cell activation, successful IFN-α14 therapy was associated with increased intrinsic and innate immunity, including significantly higher induction of tetherin and MX2, increased APOBEC3G signature mutations in HIV-1 proviral DNA, and higher frequencies of TRAIL(+) NK cells. These results identify IFN-α14 as a potent new therapeutic that operates via mechanisms distinct from those of antiretroviral drugs. The ability of IFN-α14 to reduce both viremia and proviral loads in vivo suggests that it has strong potential as a component of a cure strategy for HIV-1 infections. The broad implication of these results is that the antiviral efficacy of each individual IFN-α subtype should be evaluated against the specific virus being treated., Importance: The naturally occurring antiviral protein IFN-α2 is used to treat hepatitis viruses but has proven rather ineffective against HIV in comparison to triple therapy with the antiretroviral (ARV) drugs. Although ARVs suppress the replication of HIV, they fail to completely clear infections. Since IFN-α acts by different mechanism than ARVs and has been shown to reduce HIV proviral loads, clinical trials are under way to test whether IFN-α2 combined with ARVs might eradicate HIV-1 infections. IFN-α is actually a family of 12 distinct proteins, and each IFN-α subtype has different efficacies toward different viruses. Here, we use mice that contain a human immune system, so they can be infected with HIV. With this model, we demonstrate that while IFN-α2 is only weakly effective against HIV, IFN-α14 is extremely potent. This discovery identifies IFN-α14 as a more powerful IFN-α subtype for use in combination therapy trials aimed toward an HIV cure., (Copyright © 2016, American Society for Microbiology. All Rights Reserved.)